Cytokines like IFN-gamma and TNF-alpha have pleotrophic effects on target cells. The mechanism(s) by which these cytokines exert their biochemical modulating effects on cells has not been defined conclusively. Investigators have described the activation of second messenger systems following ligand-receptor interaction, and it has been demonstrated that ligand-receptor complexes are internalized by receptor-mediated endocytosis into endosomes and lysosomes within the cell. We considered the possibility that these cytokines, after internalization, could act intracellularly to modulate the biochemical process of the cell. This hypothesis was tested by directly introducing cytokines into the cytoplasm of the cell via microinjection. We determined that IFN-gamma and TNF-alpha do have intracellular roles in that injection of human IFN-gamma into murine macrophages induced Ia antigen expression on the cell surface and that injection of TNF-alpha induced rapid cytotoxicity, killing target cells through the activation of a cytoplasmic endonuclease. IL-1 and IL-2 were found not to have an intracellular biological role, since injection of IL-l/IL-2 did not result in the induction of DNA synthesis normally seen for the binding of these cytokines to their receptors.